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Investigation of ionic channel and its * cance in *

离子通道及其*癌症的研究*

基本信息

批准号：
11670568
负责人：
NAKAJIMA Toshiaki
金额：
$ 0.96万
依托单位：
University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11670568/
关键词：
Bronchial smooth muscle endothelin-1 ionic currents Patch champ K^+ channel Anti-serces Non-selective cation channels Biorchial asthma Brachial Smooth muscle Anti-sanse Kir2.1 Endothelin-1 Patch-clamg Intracellulor Ca^<2+> ions /

项目摘要

1) Electro-physiological effects of endothelin-1 on human bronchial smooth muscle cells : We investigated the effects of endothelin-1 and histamine on electro-physiological effects of endothelin-1 on human bronchial smooth muscle cells, by means of the patch-clamp techniques and Ca^<2+> measurements using fura-2 AM, Both endothelin-1 and histamine first increased [Ca^<2+>]_1 by the release of Ca^<2+> from store sites, then fol1owed by the sustained Ca^<2+> influx. The [Ca^<2+>]_1 increase induced by endothelin-1 was mediated by both ET_A and ET_B receptors. The activation of Ca^<2+>-permeable nonselective cation channels (I_<NS>) was involved in the sustained rise in [Ca^<2+>]_1, where ET_A and ET_B receptors were involved. The EC_<50> value for endothelin-1 to activate I_<N,S-> was approximately 10 nM.In addition, presence of both ET_A and ET_B mRNA was confirmed by RT-PCR in these cells and immunostaining of ET receptors in human airway preparations.2) K^+ channels in human bronchial smooth muscle cells : We examined the properties of K^+ channels, by using the patch clamp techniques and the RT-PCR analysis of mRNA and antisense oligonucleotide methods. In these cells, we found the voltage-dependent K^+ channels (K_v) and the inwardly rectifying K^+ channels (Kir), which contribute to forming the resting membrane potential. The Kir channels was blocked by low doses of extracellular Ba^<2+> ions with a IC_<50> of approximately - 1.8μM.RT-PCR analysis of mRNA showed transcripts for Kir2.1. but not Kir1.1, 2.2, and 2.3. Treatment of cells with antisense oligonucleotides targeted to Kir2.1 resulted in a significant decrease in current density of the Kir current. These results demonstrate that the inward rectifier K^+ current (Kir) is present in human bronchial smooth muscle cells. and the Kir2.1 gene encodes the Kir channel protein in these cells.

1)内皮素-1对人支气管平滑肌细胞的电生理效应：采用膜片钳技术和fura-2 AM测量Ca^<2+>，研究了内皮素-1和组胺对内皮素-1对人支气管平滑肌细胞电生理效应的影响。内皮素-1和组胺首先通过Ca^<2+>从储存位点释放升高[Ca^<2+>]_1，随后持续Ca^<2+>内流。内皮素-1诱导的[Ca^<2+>]_1升高是由ET_A和ET_B受体介导的。[Ca^<2+>]_1持续升高与Ca^<2+>]_1可渗透非选择性阳离子通道（I_<NS>）的激活有关，其中与ET_A和ET_B受体有关。内皮素-1激活I_<N,S->的EC_<50>值约为10 nM。此外，通过RT-PCR和人气道制剂中ET受体的免疫染色证实了这些细胞中ET_A和ET_B mRNA的存在。2)人支气管平滑肌细胞中的K^+通道：采用膜片钳技术、mRNA RT-PCR分析和反义寡核苷酸法研究了K^+通道的性质。在这些细胞中，我们发现了电压依赖的K^+通道（K_v）和向内整流的K^+通道（Kir），它们有助于形成静息膜电位。Kir通道被低剂量的胞外Ba^<2+ b>离子阻断，IC_<50>约为- 1.8μM。RT-PCR分析mRNA显示Kir2.1转录本。但Kir1.1、2.2和2.3就不一样了。以Kir2.1为靶点的反义寡核苷酸处理细胞，可显著降低Kir电流的电流密度。这些结果表明，在人支气管平滑肌细胞中存在向内整流K^+电流。Kir2.1基因在这些细胞中编码Kir通道蛋白。